Three CDK4/6 inhibitors were recently approved to treat breast cancer (BrCA) in combination regiments. Despite significant clinical activity, treatment of BrCA patients with these combinations does not result in a durable response. Hence, there is an unmet medical need to develop targeted therapeutic agents which will augment the activity of CDK4/6 inhibitors in BrCA and other CDK4/6-cyclin D-Myc dependent malignancies. A recent report demonstrated that PI3 kinase (PI3K) inhibition is critical to maintain CDK4/6 sensitivity of tumor cells. Hence, our central hypothesis to be tested in this proposal is that combined triple inhibition of CDK4/6, PI3K and BRD4 with one small molecule, SRX3177, will be more efficacious and less toxic than combining three separate inhibitors. An innovative component of our proposal is that we have developed in silico guiding models to design and then synthesized SRX317, the first small-molecule inhibitory chemotype that potently inhibits these three important targets, CDK4/6, PI-3K and BRD4 simultaneously in the same cell (preliminary results). We have demonstrated ?proof of concept? that SRX3177: 1) inhibits all three targets at nM potency, 2) inhibits all three targets in cell-based assays, 3) displays 20-fold greater efficacy in cell based assays for IC50 determination, 4) displays 40-fold less toxicity in normal cells, and most importantly 5) demonstrates efficacy but lacks detectable toxicity in vivo as compared to the high mortality rate observed in mice treated with 3 separate drugs of equal potency, e.g., Palbociclib (CDK4/6), BKM120 (PI3K) and JQ1 (BRD4). The significance of our proposal lies in our capacity to provide an optimizable promising single anticancer agent which will potently inhibit CDK4/6 Cyclin D and Myc via multiple orthogonal signaling mechanisms and inhibit survival signaling by blocking the PI3K pathway. This proposal will evaluate this approach by achieving the following aims setting the stage for phase II efforts to optimize the expected triple inhibitory lead compound(s) to a clinical candidate: Aim (Task) #1. Develop a potent CDK4/6 inhibitor with inhibitory properties against PI3K and BRD4. Approach: In silico design and modeling of 144 chemotypes to select 20 SRX3177 analogs. Aim (Task) #2. Characterize the 20 new compounds. Approach: Determine their CDK4/6, BRD4 and PI3K inhibition profiles and ADME properties. Aim (Task) #3. Demonstrate triple inhibitor is superior to use of three separate inhibitors in vivo. Approach: Compare toxicity and efficacy of triple inhibitors versus individual CDK4/6 + PI3K + BRD4 inhibitory conditions in vitro and in vivo. The significance of this research is it will greatly expand the efficacy of CDK4/6 inhibition against cyclin D/Myc dependent malignant diseases. This innovative approach attacks cancer blocking three distinct, cancer-driving orthogonal mechanisms with a single compound with greater safety in vivo challenging the one-drug one-target dogma.